Biophenics Call for Projects “Cell biology, clinical scenarios and cell-based high throughput screening” Unleash your imagination Foreword Basic Biology has been studying the world gene by gene, protein by protein. “The Times They Are a-Changin’”1 and ”massive cell biology” is a way to decipher basic mechanisms, to question cells, , to identify and validate functional networks, to identified drugs, etc… Biophenics (BFX) is a platform of “massive cell biology”, where many operations are automated: cell transfection with siRNA, immunofluorescence staining, imaging, image quantification and analysis, statistical analysis of data. Work is performed at the 96-well-format, using collections of siRNA or chemical compounds collections (see Annex 1). BFX by nature welcomes projects and this call opens the process of (easy) submission and selection. We offer the opportunity to conduct cell-based screens, using comprehensive or dedicated siRNA libraries or chemical compound libraries in association with various cellular models. Phenotypic modifications will be detected and quantified on the platform using fluorescence microscopy-based assays. Examples of possibilities offered by BFX are described in the attached PDF. What can BFX do with you for you? - Screen a collection of perturbators (see Annex 1 and 2) using one or several multiplexed cellular assays. We will be looking for siRNAs or drugs within a tested collection that will perturb your favorite phenotype. Those could include phenotypic assays for simple or complex cellular processes and pathways (e.g., proliferation, viability assays and apoptosis, membrane trafficking, cell signaling, …see PDF). Your phenotype or cell line can as well not be part of our collection, we will discuss with you the best way to make them compliant with the BFX screening process. Here is a brief outline of the assay implementation process: - Cells are grown in microscopy-compatible 96-well plates (Packard plates, which are compatible with our microscopes, robots, etc…) - They are transfected with siRNAs or exposed to drugs, - Days later, cells are fixed with aldehydes and stained with adequate primary and secondary antibodies (ideally using our automated workstation), - Images are acquired by using an automated imager (INCell1000). Cell features are quantified according to BFX settings or according to the image processing segmentation as set up and validated with the “user” (=you) before the screen. The microscope inside the InCell1000 is equipped with 5x, 10x, 20x and 40x objective. We routinely use the 10x even for small objects. - Data are processed and analyzed using validated statistical methods, outliers 1

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identified and delivered to the user. In case of siRNA screens, BFX will provide you with the sequences of siRNAs active in the screen. In case of compounds, you will have access, possibly on a collaboration basis, to the chemical molecules. What we cannot do for you - Know your phenotype better than you do, - Screen only a few compounds (siRNAs or chemicals) using multiple phenotypes. Remarks - We try, as much as possible, to multiplex immunofluorescence staining. If you are interested to track only one protein/marker by IF, we might as well test some additional phenotypes under your screening conditions, - Four different wavelengths are available on the automated imager; we routinely use DAPI/Hoechst in the first wavelength: three more are available for multiplexing. What is your commitment? You have to commit to the project at least one scientist: senior, post-doc, trained PhD student or engineer/technician (we do not have the resources to take care of untrained students). She/he will be in charge of the prerequisites (see attached doc), will perform the experiments within the BFX laboratory as well as the analysis, and take care of the follow-up. If none of our collections of siRNA (see Annex 1) is of interest to you, we will strive to find money to buy other collections. In this case, half of the collections (or at least 2 micromoles) shall stay at BFX. They will contribute to the BFX Phenotyping Fingerprinting Research Project. Results obtained with your siRNA will be shared with you. You will be charged for consumables cost. Count 1 €/ well for one labeling (primary + secondary antibody) if it is a commercial antibody. If your lab provides the antibody, costs will be reduced. We will discuss about the implementation of “your” phenotype as a standard phenotype on the platform. Publication policy This process is a collaboration, and BFX people will be co-authors on the first, maybe even second paper that comes out of your screens. Not the whole BFX team will sign but it would not be undue that two persons co-sign the first paper. Public Access to Data Policy By starting a collaboration with BFX, you accept that after a 6-month period of exclusive access for your group, data will be accessible to registered users, who will have signed an Agreement of Proper Behavior in terms of deontology. All groups of Curie and non-Curie groups selected within the frame of the Call qualify for registration. How can you apply? Attached is a document to be filled. Send it to a BFX Bio staff member. Application will be reviewed for technical aspects through a dialog with you. Then it will be submitted to our Scientific Committee. A reviewer will take care of your application. Answers should be provided within a month of transmission to the

Committee. Time-frame: none. Processing of applications will be on a basis “first come, first served”. Apply any time. BFX Bio Staff [email protected] [email protected] [email protected] [email protected] Scientific Committee See other doc. Annex 1. SiRNA available. Apart from specific collections (Amplicon chr 17, DrosoMan PIM), we have collections targeting phylogenic families: - Kinases (Dharmacon Smart Pools) and Qiagen v.2 (4siRNA/gene) - Small GTPases family (Qiagen; 2 siRNA/gene) - Kinesins family (Dharmacon Smart Pools, Qiagen (idem), Proligo (4siRNA/gene) - Exocyst subunits (Dharmacon OTP) - SNAREs (Dharmacon OTP) - V-ATPases (Dharmacon OTP) - Palmitoyl Transferases (Dharmacon OTP) Any other collections of genes/siRNA should either be provided by you or be the object of a joint purchase, and we will contribute to your grant application. . Annex 2. Chemical libraries available We plan to purchase a collection of highly diverse chemicals within the first quarter of 2008. It shall contain 10 000 to 14 000 components. Please note that such screens are time and money consuming.